We are currently investigating the specificity of the 5'-3' exonuclease function of DNA plymerase I under conditions where nick translation is (or is not) allowed. These studies are the beginning of a program to evaluate the kinetics of formation and sealing of nicks and gaps in vivo. So far, we have determined that in the absence of polymerization, the 5' exonuclease can digest an adjacent base. We have also found some conditions where polymerization occurs uncoupled to exonuclease digestin. The factors which determine the balance between these processes are being evaluated. In addition, we are continuing a structural analysis of the discontinuously sysnthesized DNA intermediates made on the cellophane disc system. We have found that these studies are technically exceedingly difficult. To evaluate the multi-enzymic steps necessary for such analyses, we now routinely include an internal standard. We hope in this way to be able to define and catalogue the 5' structure of the small Okazaki pieces found at the DNA replication fork of Escherichia coli.